IN SITU HYBRIDIZATION USING RADIOISOTOPE-LABELED PROBE FOR GFA-PROTEIN MESSENGER RNA : COMPARING WITH IMMUNOHISTOCHEMISTRY AND NORTHERN BLOT HYBRIDIZATION (<Symposium I>Molecular Cytochemistry of Nucleic Acids by In Situ Hybridization)

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In situ hybridization using radioisotope-labeled cDNA probe was performed on cow brain. The cDNA fragment for glial fibrillary acidic protein (GFA-protein) mRNA of cow was labeled with ^<32>P-dCTP or ^3H-dC TP by nick translation or random priming, and used as the probe. X-ray film macroautoradiograph of in situ hybridization using ^<32>P-labeled probe revealed that the positive signals for GFA-protein mRNA were mainly located in the white matter of cow brain. By in situ hybridization using ^3H-labeled probe, silver grains indicating existence of GFA-protein mRNA were detected on the cytoplasm of astroglia, but not on the nerve cells, vascular endothelial cells, choroid plexus and pia mater. The backgrounds were negligibly small. Specific grains were not observed on the control sections. These results were consistent with the findings of GFA-protein immunohistochcmistry and Northern blot hybridization, indicating satisfactory specificity of in situ hybridization by the protocol presented here. The critical analysis of the specificity of hybridigation-singles was thought to be the most important prerequisitc in interpreting the result of in situ hybridization.
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